PROJECT SUMMARY Influenza is a common respiratory illness that results in up to 500,000 deaths worldwide each year. Influenza A infection is often complicated by secondary bacterial infections of the lung, such as Staphylococcus aureus (SA) or Streptococcus pneumoniae. Our long-term goal is to develop novel therapeutic interventions for use in clinical settings to prevent morbidity and mortality from influenza-related secondary bacterial pneumonia. By identifying relevant cell signaling pathways, these studies have the potential to introduce novel therapeutic targets. IL-1 cytokines promote pro-inflammatory innate and adaptive immune responses. IL-33 is a newly identified cytokine in the IL-1 family that is expressed in epithelial barrier tissues and lymphoid tissues and stimulates the development of Type 2 immune cells. In addition, IL-33 can shift macrophage polarization towards an alternatively activated macrophage (M2a) phenotype. Based on our preliminary data, we hypothesize that IL-33 is essential to SA host defense and that preceding influenza A infection impairs IL-33 dependent macrophage function, resulting in attenuation of hose defense against SA and exacerbation of lung injury. Research aims: 1) To test the hypothesis that epithelial cell derived IL-33 is essential to SA host defense 2) To test the hypothesis that IL-33 promotes M2a macrophage polarization and enhances macrophage function during influenza/SA co-infection, and 3) To test the hypothesis that influenza/SA co- infection leads to acute and chronic epithelial injury, impaired lung function, and dysregulation of normal epithelial repair following influenza infection and that exogenous IL-33 can alleviate epithelial damage. The candidate's long term career goal is to become an independent NIH-funded physician scientist in the area of host defense of the lung. Immediate career development objectives include: 1) To develop expertise in immunologic, cellular, and molecular biologic techniques, 2) To develop skills to translate findings from cellular, molecular and animal studies into human disease, 3) To become proficient in the assessment of pulmonary physiology in animal models, and 4) To enhance communication and leadership skills. The proposed training plan will provide the candidate with the opportunity to expand her knowledge base to include advanced immunologic and physiologic techniques. Through direct teaching in the laboratory, coursework and conferences, the candidate will acquire advanced training in flow cytometry, in situ hybridization, RNA sequencing, use of flexiVent for lung function testing, and grant writing/presentation/leadership skills. The resources and expertise of mentors Drs. Alcorn and Kolls combined with the candidate's scientific advisory committee and the rich research environment at the University of Pittsburgh assure the candidate's successful transition to an independent investigator.